Wang Yuwen, Yuan Jianshu, Yang Liangyan, Wang Pengyun, Wang Xiajun, Wu Yue, Chen Kan, Ma Rong, Zhong Yike, Guo Xiaohong, Gong Yan, Gui Mengfang, Jin Yaming
Department of Ophthalmology, Ningbo Eye Hospital, China.
Adv Clin Exp Med. 2018 Jul;27(7):899-905. doi: 10.17219/acem/70418.
As a clinically important natural isoquinoline alkaloid, berberine has been reported to possess various pharmacological effects.
This study was aimed to investigate the effect of berberine on cell migration and invasion in human retinoblastoma (Rb) cells.
The cytotoxicity of berberine was estimated by 3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide (MTT) assay. After being stimulated with berberine under various concentrations, the cell migration and invasion were evaluated by transwell assay. Then, the expression levels of epithelial-mesenchymal transition (EMT) markers were determined by quantitative reverse transcription PCR (qRT-PCR) and western blot analysis. Furthermore, the phosphorylation levels of protein kinase B (Akt) and p38 were detected by western blot analysis. Finally, the effect of phosphatidylinositol-3-kinase (PI3K) and p38 inhibitors on cell migration and invasion was estimated by transwell assay. Untreated cells acted as control for all the experiments.
The concentrations of berberine for further studies were controlled in a range of 0 to 100 μM. The cell migration and invasion were both suppressed by berberine in a dose-dependent manner compared to the control (p < 0.05 or p < 0.001). Berberine remarkably down-regulated expression of E-cadherin and up-regulated expression of vimentin and α-SMA compared to the control (p < 0.01 or p < 0.001). Furthermore, the phosphorylation levels of Akt and p38 were both down-regulated by berberine in comparison to the control. Furthermore, the addition of berberine accompanied by LY294002 or SB203580 significantly suppressed cell migration and invasion compared to the addition of berberine alone (p < 0.05).
Berberine suppressed cell migration and invasion via inactivation of PI3K/Akt and p38.
作为一种具有临床重要性的天然异喹啉生物碱，黄连素已被报道具有多种药理作用。
本研究旨在探讨黄连素对人视网膜母细胞瘤（Rb）细胞迁移和侵袭的影响。
采用3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐（MTT）法评估黄连素的细胞毒性。在不同浓度的黄连素刺激后，通过Transwell实验评估细胞迁移和侵袭能力。然后，通过定量逆转录PCR（qRT-PCR）和蛋白质免疫印迹分析确定上皮-间质转化（EMT）标志物的表达水平。此外，通过蛋白质免疫印迹分析检测蛋白激酶B（Akt）和p38的磷酸化水平。最后，通过Transwell实验评估磷脂酰肌醇-3-激酶（PI3K）和p38抑制剂对细胞迁移和侵袭的影响。所有实验均以未处理的细胞作为对照。
进一步研究中黄连素的浓度控制在0至100μM范围内。与对照组相比，黄连素以剂量依赖性方式抑制细胞迁移和侵袭（p < 0.05或p < 0.001）。与对照组相比，黄连素显著下调E-钙黏蛋白的表达，上调波形蛋白和α-平滑肌肌动蛋白的表达（p < 0.01或p < 0.001）。此外，与对照组相比，黄连素下调Akt和p38的磷酸化水平。此外，与单独添加黄连素相比，同时添加黄连素和LY294002或SB203580显著抑制细胞迁移和侵袭（p < 0.05）。
黄连素通过使PI3K/Akt和p38失活来抑制细胞迁移和侵袭。
